PROJECT SUMMARY Polycystic ovary syndrome (PCOS) is a leading cause of female infertility and a tremendous health burden, associated with complications such as cardiovascular disease and type 2 diabetes. The etiology of PCOS is unknown, but twin studies suggest that over 70% of PCOS pathogenesis can be explained by genetics. Genome-wide association studies (GWAS) aimed to identify the genetic components of PCOS uncovered two single nucleotide polymorphisms (SNPs) in the 5? upstream regulatory region of the gene encoding the follicle- stimulating hormone (FSH) beta subunit in association with PCOS and gonadotropin levels. The gonadotropins, FSH and luteinizing hormone (LH), represent the pituitary output of the HPG axis. Women with PCOS often have an elevated LH to FSH ratio, indicating that deficient FSH secretion may play a causative role in the development of PCOS. The two PCOS-related SNPs identified through GWAS (referred to here as ?rs05? and ?rs06?) are contained within a short, evolutionarily conserved element, which is suggestive of an important biological function. We hypothesize that the conserved element functions as an enhancer of FSH?, is required for female fertility, and that the SNPs alter FSH? transcription. We will test this hypothesis through two specific aims, which will address in vitro and in vivo (1) the role of the conserved element as a FSH? transcriptional enhancer, and (2) the effect of the SNPs on FSH? transcription and the underlying molecular mechanisms. Both aims are well-supported by our preliminary in vitro data, which reveal that the conserved element from both the human and mouse genome functions as an enhancer of FSH? and demonstrates an effect of both SNPs on FSH? transcription and transcription factor binding. The proposed experiments will expand upon these data through both in vitro and in vivo experiments. In vitro, we will map enhancer and repressor sites within the conserved element, assess chromatin status near the conserved element, and identify transcription factor binding sites that are functionally altered by the SNPs. To elucidate how effects of the conserved element and SNPs interact with hormones that regulate FSH? transcription, we will perform all in vitro experiments in basal conditions and with treatment of gonadotropin-releasing hormone, activin, and testosterone. To determine how the conserved element and the SNPs affect female fertility in vivo, we will develop two novel mouse models, one with a deletion of the conserved element and one with the rs06 SNP minor allele (the rs05 SNP is not conserved between humans and mice). We will conduct a thorough fertility analysis of both mouse lines, measuring ovulation, fecundity, and hormone levels. Through our specific aims, this proposal will define novel factors that regulate FSH? transcription and will help us to understand the contributions of FSH? in the pathophysiology of PCOS, which will lead to improved diagnosis and treatment.